Biology subjects

Gerzanich, V.

Publications and source records attributed to Gerzanich, V..

8 recordsLinked to original sources

bioRxiv2025

Single-nucleus RNA sequencing reveals the cellular diversity of cerebrospinal fluid in the context of intraventricular hemorrhage

BackgroundIntraventricular hemorrhage (IVH) is a common and severe complication of hemorrhagic brain injury. Current treatments offer limited improvement in long-term neurological outcomes. Inflammatory responses in the cerebrospinal fluid (CSF) after IVH are thought to drive secondary injury, but the cellular mechanisms underlying this inflammation remain poorly defined. MethodsWe performed single-nucleus RNA sequencing of leukocytes isolated from CSF collected through external ventricular drains in subjects with intracerebral (n = 6) or subarachnoid (n = 1) hemorrhage. We characterized transcriptionally distinct subpopulations of neutrophils, monocytes, and lymphocytes by comparison to reference datasets. Cell-cell signaling networks were analyzed to infer cytokine-mediated communication, and a flow cytometry panel was developed to validate transcriptomic findings in independent CSF samples. ResultsWe obtained 11,191 high-quality nuclei comprising neutrophils (53.8%), monocytes (26.1%), lymphocytes (17.8%), and non-immune cells (2.4%). Neutrophils segregated into Nascent, Quiescent, and Interferon-Activated states. Monocytes exhibited classical phenotypes that include interferon-activated states (characterized by expression of VCAN or PROK2) and CXC-chemokine expressing states (characterized by expression of CXCL5 or CXCL8). Lymphocytes were mainly naive and central memory CD4 T cells. Cell-cell signaling analysis predicted strong CXC chemokine signaling from monocytes to neutrophil subsets and IL-1 family-driven inflammatory responses across multiple populations. Type I and III interferon signaling defined a neutrophil population not previously described in the central nervous system. ConclusionThis study delineates the diverse cellular immune landscape of CSF after IVH. Transcriptomic profiles reveal interferon, IL-1, and CXC chemokine signaling networks as potential therapeutic targets to mitigate secondary injury.

genomics↗

bioRxiv2025

Dose Escalation in Pentylenetetrazol Kindling Detects Differences in Chronic Seizure Susceptibility

ObjectivePentylenetetrazol (PTZ) kindling is a widely used model for inducing epileptogenesis and evaluating long-term seizure susceptibility differences among animals. This model is typically performed by chronic, repetitive exposures to a constant subconvulsive PTZ dose. However, the effectiveness of the commonly used subconvulsive dose (35mg/kg) varies among different animal groups and experimental conditions due to factors such as species, age, sex, and genetic background. The objective of this study was to characterize a novel model of kindling, the PTZ Dose Escalation (PTZ-DE) model, which assesses chronic seizure threshold with enhanced sensitivity by empirically determining the minimally effective dose to induce PTZ kindling for specific experimental conditions. MethodsThis study investigated the efficacy and validity of the PTZ-DE model by comparing its performance to the standard PTZ kindling approach across a series of conditions. First, the ability of the PTZ-DE model to produce the gradual increase in chronic seizure severity response characteristic of PTZ kindling was compared to the standard model across animal background characteristics (strain, sex). Next, the validity of this model was investigated by determining if the PTZ-DE model could replicate similar changes in chronic seizure susceptibility previously published using the standard approach after traumatic brain injury (TBI). Lastly, the PTZ-DE models efficacy to detect seizure differences was measured in a condition (glyburide treatment) in which alterations to chronic seizure susceptibility were not detected with standard kindling. ResultsThis study found that the PTZ-DE model corrects for background differences in PTZ susceptibility, replicates known differences in chronic seizure thresholds after TBI, and identifies new alterations in seizure threshold not detected with traditional kindling methods. SignificanceThe PTZ-DE model may prove to be a superior tool to standard PTZ kindling for discovering new pathological mechanisms of epileptogenesis and for developing targeted therapies for chronic seizure management, as evidenced by its ability to detect subtle differences in seizure susceptibility across various experimental conditions.

neuroscience↗

bioRxiv2024

RiboTag RNA Sequencing Identifies Local Translation of HSP70 In Astrocyte Endfeet After Cerebral Ischemia

Brain ischemia causes disruption in cerebral blood flow and blood-brain barrier (BBB) integrity which are normally maintained by the astrocyte endfeet. Emerging evidence points to dysregulation of the astrocyte translatome during ischemia, but its effects on the endfoot translatome are unknown. In this study, we aimed to investigate the early effects of ischemia on the astrocyte endfoot translatome in a rodent model of cerebral ischemia-reperfusion. To do so, we immunoprecipitated astrocyte-specific tagged ribosomes (RiboTag IP) from mechanically isolated brain microvessels. In mice subjected to middle cerebral artery occlusion and reperfusion and contralateral controls, we sequenced ribosome-bound RNAs from perivascular astrocyte endfeet and identified 205 genes that were differentially expressed in the translatome after ischemia. Pathways associated with the differential expressions included proteostasis, inflammation, cell cycle, and metabolism. Transcription factors whose targets were enriched amongst upregulated translating genes included HSF1, the master regulator of the heat shock response. The most highly upregulated genes in the translatome were HSF1-dependent Hspa1a and Hspa1b, which encode the inducible HSP70. We found that HSP70 is upregulated in astrocyte endfeet after ischemia, coinciding with an increase in ubiquitination across the proteome. These findings suggest a robust proteostasis response to proteotoxic stress in the endfoot translatome after ischemia. Modulating proteostasis in endfeet may be a strategy to preserve endfeet function and BBB integrity after ischemic stroke.

neuroscience↗

bioRxiv2023

Region-specific homeostatic identity of astrocytes is essential for defining their reactive phenotypes following pathological insults

The transformation of astrocytes into reactive states constitutes a biological response of the central nervous system under a variety of pathological insults. Astrocytes display diverse homeostatic identities, which are developmentally predetermined and regionally specified. Upon transformation into reactive states associated with neurodegenerative diseases and other neurological disorders, astrocytes acquire diverse reactive phenotypes. However, it is not clear whether their reactive phenotypes are dictated by regionspecific homeostatic identity or, alternatively, by the nature of an insult. To address this question, regionspecific gene expression profiling was performed for four brain regions (cortex, hippocampus, thalamus and hypothalamus) in mice using a custom Nanostring panel consisting of selected sets of genes that report on astrocyte functions and their reactivity for five conditions: prion disease, traumatic brain injury, brain ischemia, 5XFAD Alzheimers disease model and normal aging. Upon transformation into reactive states, genes that are associated predominantly with astrocytes were found to preserve region-specific signatures suggesting that they respond to insults in a region-specific manner. A common gene set was found to be involved in astrocyte remodeling across insults and normal aging. Regardless of the nature of an insult or insult-specificity of astrocyte response, strong correlations between the degree of astrocyte reactivity and perturbations in their homeostasis-associated genes were observed within each individual brain region. The insult-specific populations did not separate well from each other and instead partially overlapped, forming continuums of phenotypes. The current study demonstrates that astrocytes acquire their reactive phenotypes according to their region-specific homeostatic identities. Within these region-specified identities, reactive phenotypes show continuums of states, partially overlapping between individual insults.

neuroscience↗

bioRxiv2021

Traumatic brain injury alters dendritic cell differentiation and distribution in lymphoid and non-lymphoid organs

Pathophysiological consequences of traumatic brain injury (TBI) mediated secondary injury remain incompletely understood. In particular, the impact of TBI on the differentiation and maintenance of dendritic cells (DCs), remains completely unknown. Here, we report that DC-differentiation, maintenance and functions are altered at both early and late phases of TBI. Our studies identify that; 1. frequencies and absolute numbers of DCs in the spleen and BM are altered at both acute and late phases of TBI; 2. surface expression of key molecules involved in antigen presentation of DCs were affected both at early and late phases of TBI; 3. distribution and functions of tissue-specific DC subsets of both circulatory and lymphatic systems were imbalanced following TBI; 4. early differentiation program of DCs, especially the commitment of hematopoietic stem cells to common DC progenitors, were deregulated after TBI; and 5. intracellular ROS levels were reduced in DC progenitors and differentiated DCs at both early and late phases of TBI. Our data demonstrate, for the first time, that TBI affects the distribution pattern of DCs and induces an imbalance among DC subsets in both lymphoid and non-lymphoid organs. In addition, the current study demonstrates that TBI results in reduced levels of ROS in DCs at both early and late phases of TBI, which may explain altered DC differentiation paradigm following TBI. A deeper understanding on the molecular mechanisms that contribute to DC defects following TBI would be essential and beneficial in treating infections in patients with acute central nervous system (CNS) injuries.

immunology↗

bioRxiv2021

Genome-wide characterization of SARS-CoV-2 cytopathogenic proteins in the search of antiviral targets

Therapeutic inhibition of critical viral functions is important for curtailing coronavirus disease-2019 (COVID-19). We sought to identify antiviral targets through genome-wide characterization of SARS-CoV-2 proteins that are crucial for viral pathogenesis and that cause harmful cytopathic effects. All twenty-nine viral proteins were tested in a fission yeast cell-based system using inducible gene expression. Twelve proteins including eight non-structural proteins (NSP1, NSP3, NSP4, NSP5, NSP6, NSP13, NSP14 and NSP15) and four accessory proteins (ORF3a, ORF6, ORF7a and ORF7b) were identified that altered cellular proliferation and integrity, and induced cell death. Cell death correlated with the activation of cellular oxidative stress. Of the twelve proteins, ORF3a was chosen for further study in mammalian cells. In human pulmonary and kidney epithelial cells, ORF3a induced cellular oxidative stress associated with apoptosis and necrosis, and caused activation of pro-inflammatory response with production of the cytokines TNF-, IL-6, and IFN-{beta}1, possibly through the activation of NF-{kappa}B. To further characterize the mechanism, we tested a natural ORF3a Beta variant, Q57H, and a mutant with deletion of the highly conserved residue, {Delta}G188. Compared to wild type ORF3a, the {Delta}G188 variant yielded more robust activation of cellular oxidative stress, cell death, and innate immune response. Since cellular oxidative stress and inflammation contribute to cell death and tissue damage linked to the severity of COVID-19, our findings suggest that ORF3a is a promising, novel therapeutic target against COVID-19. SignificanceThe ongoing SARS-CoV-2 pandemic has claimed over 5 million lives with more than 250 million people infected world-wide. While vaccines are effective, the emergence of new viral variants could jeopardize vaccine protection. Antiviral drugs provide an alternative to battle against COVID-19. Our goal was to identify viral therapeutic targets that can be used in antiviral drug discovery. Utilizing a genome-wide functional analysis in a fission yeast cell-based system, we identified twelve viral candidates, including ORF3a, which cause cellular oxidative stress, inflammation and apoptosis and necrosis that contribute to COVID-19. Our findings indicate that antiviral agents targeting ORF3a could greatly impact COVID-19.

molecular biology↗

bioRxiv2021

Immunomodulatory Therapy with Glatiramer Acetate Reduces Endoplasmic Reticulum Stress and Mitochondrial Dysfunction in Experimental Autoimmune Encephalomyelitis

Endoplasmic reticulum (ER) stress and mitochondrial dysfunction are found in lesions of multiple sclerosis (MS) and of experimental autoimmune encephalomyelitis (EAE), an animal model of MS, and may contribute to the neuronal loss that underlies permanent impairment. These pathological changes are due to the neuroinflammation that characterizes MS and the strong interplay between the ER and mitochondria. We investigated whether immunomodulatory drug glatiramer acetate (GA) can reduce these changes in the spinal cords of chronic EAE mice by using routine histology, immunostaining, and electron microscopy. EAE spinal cord tissue exhibited increased infiltration/inflammation (upregulation of proinflammatory cytokines), demyelination, mitochondrial dysfunction (increased fission, decreased fusion, and increased biogenesis), ER stress, downregulation of NAD+ dependent pathways, and increased neuronal death. GA reversed these pathological changes, suggesting that immunomodulating therapy can attenuate ER stress, mitochondrial dysfunction, apoptotic cell death, and demyelination in the CNS and increase NAD+ utilizing activities by suppressing neuroinflammation.

neuroscience↗

bioRxiv2020

HIV-1 Vpr-induced Proinflammatory Response and Apoptosis are Mediated through the Sur1-Trpm4 Channel in Astrocytes

There are about 38 million people currently living with HIV/AIDS worldwide. Successful treatment with combinational antiretroviral therapies (cART) can eliminate active replicating viruses and prolong lives to nearly normal lifespans. However, the new challenge faced by more than half of those HIV-infected and aging patients is chronic CNS neuroinflammation, which leads to HIV-associated neurocognitive disorders (HAND). While severe and progressive HAND has decreased significantly due to cART, chronic HAND often persists, resulting in high rates of delirium, dementia and depression that could lead to suicide. Indeed, the risk of suicide mortality in HIV-infected persons is significantly higher than in HIV-uninfected counterparts. Nevertheless, the mechanism of neuropathogenesis underlying HAND is not well understood. HAND is typically characterized by HIV-mediated glial neuroinflammation and neurotoxicity. Interestingly, the severity of some HAND does not always correlate with the levels of HIV, but rather with glial activation, suggesting other HIV-associated factors, not the whole virus per se, contribute to those HAND. HIV-1 viral protein R (Vpr) might be one of those viral factors, because Vpr induces neuroinflammation and causes neuronal apoptosis. The objective of this study was to delineate the specific role(s) of Vpr in activation of host neuroinflammation and neurotoxicity, as well as its contribution to HAND. In this report, we show correlations between HIV expression and activation of proinflammatory markers (TLR4, TNF, and NF{kappa}B) and the Sur1-Trpm4 channel in astrocytes of HIV-infected postmortem human and transgenic mouse brain tissues. We further show that Vpr alone activate the same set of proinflammatory markers in an astrocytic cell line SNB19. Vpr-induced host cell proinflammatory responses result in apoptotic cell death. Together, our data suggest that HIV-1 Vpr-induced proinflammatory response and apoptotic cell death are mediated through the Sur1-Trpm4 channel in astrocytes.

neuroscience↗